WO2001027586A1 - Methode de preparation d'un echantillon de tissu biologique et instrument et dispositif afferent - Google Patents

Methode de preparation d'un echantillon de tissu biologique et instrument et dispositif afferent Download PDF

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Publication number
WO2001027586A1
WO2001027586A1 PCT/JP2000/003107 JP0003107W WO0127586A1 WO 2001027586 A1 WO2001027586 A1 WO 2001027586A1 JP 0003107 W JP0003107 W JP 0003107W WO 0127586 A1 WO0127586 A1 WO 0127586A1
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WIPO (PCT)
Prior art keywords
blade
blades
biological tissue
sample
sliced
Prior art date
Application number
PCT/JP2000/003107
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English (en)
Japanese (ja)
Inventor
Akiyo Shigematsu
Yuko Hamai
Naomi Motoji
Original Assignee
Institute Of Whole Body Metabolism
Japan Science And Technology Corporation
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Application filed by Institute Of Whole Body Metabolism, Japan Science And Technology Corporation filed Critical Institute Of Whole Body Metabolism
Publication of WO2001027586A1 publication Critical patent/WO2001027586A1/fr

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
    • G01N1/286—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q involving mechanical work, e.g. chopping, disintegrating, compacting, homogenising
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N1/00—Sampling; Preparing specimens for investigation
    • G01N1/02—Devices for withdrawing samples
    • G01N1/04—Devices for withdrawing samples in the solid state, e.g. by cutting
    • G01N1/06—Devices for withdrawing samples in the solid state, e.g. by cutting providing a thin slice, e.g. microtome

Definitions

  • the present invention relates to a method for preparing a biological tissue slice sample, and an instrument and an apparatus used for the method.
  • the present invention relates to a method for preparing a biological tissue slice sample in which cells are kept at least morphologically stable, and an instrument and an apparatus used for the method.
  • a combined blade in which four or more plate-shaped blades having the same straight or curved blade are fixed at predetermined intervals in parallel with each other and the blades are also parallel.
  • the biological tissue block is cut into a predetermined thickness by using the above method, and a biological tissue slice is obtained.
  • Japanese Patent Application Laid-Open No. Hei 7-218398 discloses a method of inserting biological tissue slices from the gap of the blade by inserting the biological tissue slices into the gaps of the blade. By this method, a sliced sample can be prepared from a biological tissue kept near the freezing point.
  • tissue section was approximately 0.1 mm from the cut section.
  • the cells must be at least morphologically stable, that is, the microstructure and the semipermeable membrane structure should be in a normal state without causing cell death and nucleus enrichment.
  • the present inventors have studied a method for morphologically maintaining cells stably without causing cell death as well as nucleus enrichment. Produces slices of thickness. 2) Makes the number of times of friction between the surface corresponding to the entire cut surface and the blade one. 3) Comb-shaped (plastic) plate to release sliced sections from multiple blades. 4) By covering the side walls of both blades with a coating of a material with excellent insulation properties, it is possible to prevent the occurrence of cross-sectional injury current during cutting of sliced slices.5) Furthermore, to prevent the generation of injury current, making sexual cold physiological salt solution 1 0- 3 mm ol added Tran Kiraiza one to a concentration liver tissue slices in immersed state and 6) slice section thickness required such that a 2 mm thick Admitted. The present invention has been made based on these findings.
  • An object of the present invention is to realize a method for preparing a sliced biological tissue sample in which cells are kept at least morphologically stable.
  • Still another object of the present invention is to realize an apparatus for rapidly preparing a biological tissue slice sample under a constant temperature and sterile atmosphere. Disclosure of the invention
  • the above object of the present invention is the same straight or curved shape made of ceramic
  • the biological tissue block is set to a specified thickness by using a combined blade that holds two or more plate-shaped blades that have the same blade and that are parallel to each other at predetermined intervals and the blades are also parallel.
  • a biological tissue sample obtained by cutting and obtaining biological tissue slices, inserting extrusion members of a predetermined thickness that can be inserted between the blades into gaps between the blades, and extruding the biological tissue slices between the blades. This was achieved by a fabrication method.
  • the object of the present invention is also to provide two or more plate-like blades made of ceramic and having the same straight or curved blade, and these blades are parallel to each other and the blades are also parallel. And a pushing member having a predetermined thickness that can be inserted between the blades, so that the blades are held at predetermined intervals so that the pushing members can be inserted therebetween.
  • the pushing members are held parallel to each other (including fixing), and the pushing members are held at predetermined intervals so that they can be smoothly inserted between the blades, so that the pushing members are inserted between the blades, respectively.
  • the blade 1 is made of ceramic. When metal is used, a wound current is generated near the cut cells, and the cells in the area where the wound current leaks are damaged and lose cell stability. On the other hand, in the case of a ceramic blade, no injury current is generated, and the stability of the cut cells is increased.
  • Zirconia ceramic or the like is suitable as the ceramic constituting the plate-shaped blade.
  • the number of plate-shaped blades may be any number of two or more. However, a certain number of large number of blades S (for example, four blades) can be obtained with a uniform thickness.
  • the size of the blade depends on the size of the object to be sliced and is not particularly limited. Force Blade width (perpendicular to blade) is 2 to 15 mm, blade length is 10 to 0 mm.
  • the thickness of the blade is usually between 05 and lmm, but preferably less than 0.4 mm.
  • the minimum thickness of the blade is determined by its mechanical strength and is usually limited to 0.05 mm.
  • Blade may be attached only on one side of the blade may be subjected to double-sided c
  • the shape of the blade may be a straight line or a curved line (for example, an arc), but is preferably an arc. Further, it is preferable to hold the cutting tool so that it can rotate about a common axis perpendicular to the surface, so that the cutting tool having the arc-shaped blade rotates along the surface.
  • the blade is formed in a disk shape having a blade on a circumference and rotated around a common axis.
  • the rotation may be reciprocating, but may be continuous or intermittent in one direction, and a rotation speed of about 10 to 60 rotations per minute is appropriate.
  • the blade of each blade is located in one plane perpendicular to the blade, but the cutting edge may be displaced in the direction of the blade surface (gooseling).
  • the spacing between the blades depends on the desired thickness of the slices, but is usually between 0.2 and 5 mm. If the tissue is particularly soft, it is necessary to widen the interstices between the blades. For example, in the case of the brain, it is preferable that the distance be 0.8 mm or more.
  • the extruding members must each be easily inserted between the blades and be able to extrude the flakes without causing any significant deformation :: Therefore, the thickness is slightly less than the spacing between the blades, but usually 0.15.
  • the width of the extruded member which is 4.9 mm, is also smaller than the blade length of the blade. It is always 8 to 250mm.
  • the length of the extruded member (perpendicular to the blade edge) is slightly larger than the blade width, usually 5 to 60 mm, so that the flakes can be completely extruded.
  • the extruded member should be made of a material that will not be damaged or corroded by the tissue material and will not affect the biological properties or function of the tissue.
  • the blade holding member and the pushing member holding member may come into contact with the tissue material, it is preferable to use the same material.
  • the flakes can be extruded from the blade edge or from the opposite side of the blade. When extruding a sample slice from the direction of the blade, the slice will pass between the blades. This is advantageous if the blade is disc-shaped.
  • the cutting tool may be fixed and the extruding member may be moved.However, especially when the cutting tool has a disk shape, the extruding member is fixed, the cutting tool also functions as the cutting tool, and the cutting tool is moved to perform cutting and pushing. Moore, c
  • the biological tissue thin section prepared by the blade is extruded from between the blades by an extruding member and floated in cold water. It is preferable to perform the operations of slicing and extruding while immersing the combined knife in cold water and keeping the temperature around the instrument at a low temperature with ice, a cold insulator or the like.
  • the method and device of the present invention are capable of obtaining a large number of bioactive tissue slices from a tissue block in a short time without being affected by cutting.
  • a combined blade in which two or more plate-shaped blades made of ceramic and having the same straight or curved blade are fixed at predetermined intervals in parallel with each other and the blades are also parallel. Is used.
  • tissue block When the tissue block is used to cut through the tissue block, a plurality of tissue slices corresponding to the number of blades are cut out of the block at once, with a thickness corresponding to the blade interval, and the slices are once located between the blades.
  • Extruding members having a thickness that can be inserted between the blades and fixed parallel to each other can be inserted between the blades.
  • the extruding member When the extruding member is inserted between the blades, the flakes between the blades are extruded.
  • the ejected flakes have a thickness approximately equal to the spacing of the blades. That is, a tissue slice having a thickness corresponding to the interval between the blades is obtained.
  • the tissue slices have near normal bioactivity with less damage to the approximately 100 micron layer near the cut surface of the cut tissue sample. This is because when the blade is made of a good electrical conductor such as stainless steel, the positive potential of the cells on the cut surface is lost, whereas when the blade is made of ceramic, this potential is maintained. There will be.
  • the biological tissue sliced sample preparation device of the present invention includes a constant temperature bath containing the sample preparation device main body, a sterilization lamp disposed in the constant temperature bath to sterilize the inside of the constant temperature bath, and connected to the constant temperature bath and has a refrigerant therein.
  • a cooler for circulating water sterilization and slicing work can be performed in an atmosphere maintained at a low temperature.
  • a tray provided with a heat exchanger connected to a refrigerant circulated from a cooler is provided in a constant temperature bath, and a vat installed in the tray and filled with a buffer solution is provided.
  • the buffer agitating means by disposing the buffer agitating means, the temperature measuring means, and the sample holding means in the vat, it is possible to stabilize the internal temperature and temporarily store the sample in that state.
  • the main body of the sample preparation apparatus in the present invention is fixedly installed in a thermostat.
  • a fixed guide provided with a guide groove along the longitudinal direction, a slide block for holding a biological tissue and being relatively movable along the guide groove of the fixed guide, and standing at one end of the fixed guide.
  • a bearing bracket rotatably supporting a plurality of disk-shaped blades, and a motor disposed on the bearing bracket and rotating the disk-shaped blades via power transmission means. This enables the slicing operation to be performed quickly 5 .
  • the main body of the sample preparation apparatus includes a lid detachably attached to the upper surface of the slide block, and the lid and the slide block are provided with a groove having an interval and a number of grooves along the longitudinal direction corresponding to an arrangement interval of the disk-shaped blades.
  • FIG. 1 is a plan view, a front view, and a cross-sectional view of a combined cutter used in the present invention, wherein (A), (B), and (C) are.
  • FIGS. 2 (A), (B) and (C) are elevation views showing a cutting procedure using the biological tissue sliced sample preparation device according to the first embodiment of the present invention.
  • Fig. 3 is a plan view of the device for preparing a biological tissue sliced sample.
  • Fig. 4 is a side view of the biological tissue slice preparation device.
  • FIG. 5 shows (A), (B) and (C) biological tissues according to the second embodiment of the present invention. It is an elevation view showing a cutting procedure using a sliced sample preparation instrument.
  • FIG. 6 shows (A), (B) and (C) a plan view, a side view and a front view of the thermostatic oven according to the present invention.
  • FIG. 7 is a perspective view of the same.
  • FIG. 8 is a plan view, a front view, and a partially cutaway side view of the heat exchanger and the bat according to the present invention, (A), (B), and (C).
  • FIGS. 9 (A) and (B) are a plan view and a left side view showing a specific configuration example of the sliced sample manufacturing apparatus according to the present invention.
  • FIG. 11 is a perspective view of the same device.
  • FIG. 12 is a ⁇ ⁇ ⁇ tissue stained image of a liver slice using a ceramic slicer.
  • ⁇ immediately after slice production,
  • ⁇ ⁇ ⁇ ⁇ after 1 hour culture,
  • C after 4 hour culture,
  • D after 24 hour culture.
  • FIG. 13 is a ⁇ . ⁇ tissue-stained image of a liver slice using a conventional metal slicer.
  • ⁇ shows the results immediately after slice production
  • ⁇ shows the results after 1 hour of culture
  • C shows the results after 4 hours of culture
  • D shows the results after 24 hours of culture.
  • FIG. 1 shows a combined knife used in the method of the present invention.
  • FIG. 1 ( ⁇ ) is a plan view
  • ( ⁇ ) is a front view
  • FIG. 1 (C) is a cross-sectional view taken along the section line X--X.
  • the combined blade is composed of five blades 1 each having the same linear blade and a blade holding member 3 for fixing the blades 1 in parallel with each other.
  • An extruding plate 2 is provided between the blades 1, and the extruding plate 2 is fixed in parallel to each other by an extruding plate holding member 4.
  • Blade I The blades are fixed by a blade holding member 3 at regular intervals in parallel with each other, and the blades are also parallel.
  • the five blades 1 connected by the blade holding member 3 constitute a combined blade.
  • the space between the blade 1 and the push plate 2 is such that the push plate 2 is inserted into the blade 1 while being in contact with the blade 1.
  • the extruded plate 2 inserted into the gap of the blade 1 can move in the direction of the arrow in the figure, and reaches the position shown by the chain line in the figure.
  • Blade 1 is made of 0.25 mm thick zirconia ceramic.
  • the extruded plate 2 is made of plastic with a thickness of about .9 mm.
  • the thickness of the blade holding member 3 and, accordingly, the gap between the blades 1 is about 2 mm.
  • tissue slice When a biological tissue block is cut off using a combined blade consisting of blade 1 and blade fixing member 3, a plurality of tissue slices corresponding to the number of blades 1 are cut out of the block at once with a thickness corresponding to the spacing between blades 1. .
  • the flakes are once located in the gap between the blades 1.
  • the extruding plate 2 is inserted and pushed in the direction of the arrow in the figure, the flakes are pushed out of the combined blade. Its thickness is approximately equal to the spacing of the blades 1.
  • four tissue slices having a thickness corresponding to the interval between blades 1 can be obtained at once. If the rest of the block is cut in a similar manner, many tissue slices can be obtained in a short time. The obtained tissue slice retains biological activity including the surface layer.
  • FIG. 2, FIG. 3, and FIG. 4 show a first embodiment of the biological tissue sliced sample preparation device according to the present invention.
  • FIG. 2 (A), (B) and (C) are elevation views
  • FIG. 3 is a plan view in the state of FIG. 2 (A)
  • FIG. 4 is a side view.
  • the combined blade is composed of five disk-shaped blades 11 each having a blade on the circumference, and a blade support shaft 13 that holds the disk-shaped blades 11 in a mutually parallel and rotatable manner.
  • Disc shaped blade 1 1 The five disk-shaped blades 11 fixed parallel to each other at regular intervals by a blade support shaft 13 and connected by the blade support shaft 13 constitute a combined blade.
  • the sample carrier 12 for fixing the biological tissue sample to be cut has a flat surface ⁇ “12a, 12c, 12e and protrusions 12b, 12d, and the protrusions 12b, 12d and the flat portion 12c have a disc shape.
  • Parallel grooves 14 are provided at intervals so that the blade 11 can pass through.
  • the flat portion 12c between the protrusions 12b and 12d forms a recess for fixing a tissue sample, and the protrusions in 2b, 12d and flat
  • the portion between the grooves 4 of c 12c is inserted with a slight gap between the disk-shaped blades 11.
  • the sample carrier 12 is moved by the disk-shaped blades 11 through the grooves 14 as indicated by arrows in the figure. Can move in the direction of.
  • the blade 11 is made of 0.25 mm thick zirconia ceramic, and has blades on both sides on the circumference.
  • the sample carrier 12 is made of plastic.
  • the gap between the disc-shaped blades 11 is about 2 mm.
  • the tissue thickness will be approximately 2 mm.
  • a thickness of sliced into 2 mm e This nutrient broken blood vessel when the thickness is more than 2 mm, the oxygen is not supplied to the cell, also the thickness of conventional At 0.3 mm, there is a problem that the number of dead cells increases.
  • the biological tissue block is fixed in the recesses fixed to the protrusions 12b and 12d and the flat portion 12c of the sample transfer table 12, and the sample is transferred while rotating the disk blade 11 supported by the blade support shaft 13 clockwise.
  • the platform 12 is moved in the direction of the arrow at a constant speed to push down the biological tissue block. It is cut at once.
  • the tissue slice is once located in the gap of blade ⁇ 1
  • the protrusion 12d pushes the blade 11 out of the circumference of the blade 11.
  • Its thickness is substantially equal to the distance between the disc-shaped blades 11. In this way, when five blades are used, four tissue slices having a thickness corresponding to the interval between the blades can be obtained at once. If a new block is cut in the same manner, a large number of tissue slices that retain biological activity, including the surface layer, can be obtained in a short time.
  • FIG. 5 shows a second embodiment of the biological tissue sliced sample preparation device used in the present invention
  • FIGS. 5 (A), (B) and (C) are plan views in three states, respectively.
  • the structure of the combined blade is the same as that of FIG. 2, except that the disk-shaped blade 21 can move at a predetermined speed in the direction of the arrow along the sample fixture 22 while rotating. Only the others are the same.
  • the sample fixing table 22 for fixing a biological tissue sample to be cut has flat surfaces 22a, 22c, 22e and protrusions "22b, 22d, 24", similar to the general sample carrier 12.
  • the flat portion 22c between b and 22d forms a concave portion for fixing the tissue sample.
  • the disc-shaped blade 21 has a projection 22b surrounding the round portion and a groove formed in the flat portion 22c. At 24, it is inserted with a small gap.
  • the biological tissue block is fixed in the concave portion surrounded by the protruding portions 22b and 22d and the flat portion 22c of the sample fixing table 22, and the disk blade 21 supported by the blade supporting shaft 23 is rotated clockwise while rotating.
  • a plurality of tissue slices corresponding to the number of blades 21 are cut from the block at once, with a thickness corresponding to the interval of the blade 2].
  • the tissue slice is once located in the gap between the blades 21, but after the disk-shaped blade 21 moves further (in the direction of the arrow in the drawing), it stays in the concave portion of the sample fixing table 22.
  • the thickness is the interval of the disk-shaped blade 21 Almost equal. Thus, when five blades are used, four tissue slices having a thickness corresponding to the blade interval can be obtained at once. If a new block is cut in the same manner, a large number of tissue slices that retain biological activity, including the surface layer, can be obtained in a short time.
  • the embodiment described above is a general example of a method and a device. However, in order to actually implement this method, it is necessary to perform the method quickly at a constant temperature and in a sterilized atmosphere. Therefore, in the present invention, a specific device suitable for implementation has been developed.
  • FIGS. 6 to 11 show an embodiment of the apparatus.
  • FIGS. 6 and 7 show the overall configuration of a thermostatic bath for maintaining a constant temperature and sterilizing atmosphere
  • FIG. The bat placed inside and the heat exchanger in which the bat is installed are shown.
  • Figs. 9 to 11 show the main body of the sample preparation apparatus used by being installed in the bat.
  • the thermostat 100 has a portable support frame 102 with casters arranged at the lower end, and a frame 102. And a cooler 106 installed on a support plate that is inside the frame 102 and extends below it, which is a constant temperature bath main body that is installed on the top plate surface. I have.
  • the chamber 104 is a closed box made of a synthetic resin with an inclined front surface.
  • Transparent glass 108 such as acrylic glass is placed on both sides of the closed box, and acrylic glass or the like is also attached to the inclined front surface.
  • the door 110 made of transparent glass so that it can be opened and closed, the three surfaces are made transparent, facilitating external observation, and at the same time, opening and closing the door 110 enables the sample to be inserted and removed from the front. I have.
  • working rubber gloves 112 are hermetically attached, and a pair of windows 114 projecting toward the inside of the tank 104 are opened. Can be handled without directly touching the sample.
  • a germicidal lamp 116 for both sterilization and illumination is arranged at the upper part inside the chamber 104, and a rear side is connected to a refrigerant circulation tube 118 from the cooler 106 to circulate the refrigerant inside.
  • a pair of joints 120 are provided.
  • the refrigerant circulation tube 118 is connected to a heat exchanger 124 provided with a vat 22.
  • FIG. 8 shows the detailed structure of the heat exchanger 124 and the vat 122 disposed therein.
  • the heat exchanger 124 has a flat closed container shape, and has a heat exchanger body 128 having a pair of nipples 126 constituting a coolant supply / discharge boat projecting from the back, and a bat integrated at an upper part of the body.
  • a support tray 130 is provided, and a bat 122 is installed inside the tray support tray 130 and is laid.
  • the vat 122 has a stainless steel surface lined with Teflon or the like.
  • a buffer solution is filled in the inside of the vat 122, and a clamp support rail 132 is arranged at the rear of a tray 130 on which a sample preparation apparatus body described later is installed.
  • a stirring motor 138, a thermometer display section 140, and a sample holding rod 142 are mounted on the table 132 via a plurality of clamp rods 134 and clamp blocks 136, respectively.
  • a stirrer 142 hangs down at the tip of the motor 138, a sensor 144 hangs down at the bottom of the display unit 140, and a sample holder 148 with a net is fixed at the tip of the rod 14.
  • the lower end is immersed in a buffer solution stretched inside the bat 122 on the rear side thereof.
  • thermometer is not limited to a mere display, and the temperature of the cooler 106 may be controlled via a control unit by a sensor output of the thermometer.
  • the above-described sample production apparatus main body 200 is installed in a form immersed in a buffer solution in the middle.
  • sample preparation apparatus main body 200 The same sample preparation apparatus as that described in FIGS. 1 to 4 can be used for the sample preparation apparatus main body 200, and the sample preparation apparatus main body described below can also be used.
  • sample preparation instruments adopt a more specific configuration for assembling a tissue sample preparation apparatus.
  • FIGS. 9 (A), (B), FIGS. 10 (A), (B) and FIG. 11 show the specific structures.
  • a device 200 is fixed on a base 201 and has a guide groove formed along the longitudinal direction of the upper surface of the fixed guide 202, and engages with the groove of the fixed guide 202 to slide in the longitudinal direction.
  • a bracket 210 which is erected and bears the shafts of a large number of disc-shaped blades 208 as described above, a motor support frame 212 fixed to one side of the bracket 210, and a motor 214 mounted on the motor support frame 212
  • the output shaft of the motor 214 is connected to the gear 216, and the rotation is transmitted to the drive gear 220 provided at the shaft end of the blade 208 via the intermediate gear 218 :
  • the material, thickness, and arrangement interval of the blade 208 are the same as those in the above-described embodiments, and thus description thereof will be omitted.
  • a concave portion is formed on the upper surface of the slide block 204, and a concave and convex portion is formed on the bottom surface thereof.
  • Grooves 222 are formed in the convex portion along the longitudinal direction according to the arrangement interval of the blades 208, and the lid 206 is also formed. Similarly, a groove 222 is formed.
  • the lid 206 functions as a danger prevention guard, and also prevents the sample strips cut by the blade 208 from adhering to the blade 208.
  • the tip of the lid 206 has a knurling force on the outer periphery.
  • Slide block 204 A connecting knob 224 is attached, and the knob 224 is screwed into the slide block 204 side, so that the two are integrally connected.
  • the method of preparing a biological tissue sample having the above configuration is entirely performed in the chamber 102 kept in a constant temperature sterilized atmosphere.
  • the procedure for setting the sample in the chamber 102 is as follows. First, the biological tissue block is placed in the slide block 204 with the lid 206 of the manufacturing apparatus main body 200 opened, and then the lid 206 is placed on the slide block 204. The cover 206 is fixed to the slide block 204 by screwing the knob 2 24.
  • the blade 208 is rotated by turning on a switch (not shown) and driving the motor 214, so that the slide block 204 is moved by pushing the slide block 204 in the direction of the arrow in FIG. Is cut into strips of the dimensions corresponding to the arrangement interval I of the blade 208. Thereafter, the slide block 204 is pressed off or returned to its original position, and the lid 206 is opened to obtain a sample.
  • the sample preparation apparatus main body 200 by the action of the lid 2 06 arranged on the slide block 204, the strip does not adhere to the blade, it is not necessary to push the tissue strip c i.e., sliced tissue debris samples Is held in the slide block 204.
  • the lid 206 serves to prevent danger, but may have a function of holding the sliced tissue Itoda piece sample in the slide block 204.
  • the detachable slice retaining plate can provide the function of holding the sliced sample in the slide block. That is, an upper lid plate provided with a passage groove so as to overlap with a passage groove provided in the longitudinal direction of the slide block and not obstructing the passage of the round blade is attached to the slide block, and the upper end plate of the plate at the front end in the longitudinal direction is attached. Detachable so that it can be opened and closed as a top lid A retaining plate connected by a hinge.
  • the liver slice is cut by the round blade and moves up and down while the liver tissue block is cut by the round blade while keeping close contact with the gap between adjacent round blades. Without this, the round blade passes through the liver tissue block.
  • the survival of the cells prevents the cut slice from moving within the tissue mass after passing through the round blade by the function of the slice retention plate, and still contains physiological saline even after passing through the tissue blocking force of the round blade. It was found that resting in buffer was important. That is, it is necessary not only to not use stainless steel or the like which is a source of damage current after cutting but also to make the slice after cutting still.
  • the sample is cut by moving the slide block 204 by manual operation. However, it is needless to say that the sample can be replaced with an automatic feed mechanism.
  • the bracket 210 can be moved along the fixed guide 202, and in this case, either a manual operation or an automatic feed mechanism can be adopted.
  • tissue slices thickness of rat brain using thickness 0. 25 m m of zirconyl two A ceramic blade Yore , first view of the instrument of sales Enuti one Corporation 2 mm.
  • optical microscope sections were prepared by a conventional method and observed with a microscope. The cells were found to be 0.1 mm deep from the cut surface. There were only a few sites where pyknosis was observed. No abnormalities were observed in the deep cell images.
  • a zirconia ceramic blade (NTT Corporation) having a radius of 30 mm and a thickness of 0.25 mm is used. ), (B) and (C) were used to prepare rat brain tissue slices (thickness mm). After placing the slices in Xrebs-Ringer buffer at a temperature of 20 ° C for 20 minutes, optical microscope sections were prepared by a conventional method, and observed under a microscope, and the sections were cut to a depth of 0.1 mm. Nuclear enrichment was observed in fewer cells than in Example 1.
  • Healthy rats (SD, male, 7-week-old) were lightly anesthetized with ethyl ether and laparotomy was performed.
  • the rod-shaped liver tissue mass was quickly placed on the stage in cold Slices were prepared using the material preparation apparatus shown in the figure.
  • the prepared liver tissue sections were cultured in a culture solution, and tissue specimens were prepared immediately after slice preparation, 1, 4, and 24 hours later.
  • a 0.3 mm thick stainless steel razor was used as the blade 1 in FIG. 1, and a tissue slice (2 mm thick) of a rat brain was prepared using the instrument in FIG. After placing the slices in Krebs-Rhger buffer at a temperature of 20 ° C for 20 minutes, optical microscope sections were prepared by a conventional method and observed under a microscope. Nucleic enrichment was observed in most cells. All cells were dead after 60 minutes in buffer. Comparative Example 2
  • the morphological thus, a biological tissue slice sample in which cells are kept sufficiently stable can be prepared.
  • a biological tissue sliced sample in which cells are kept morphologically stable can be prepared.
  • the use of the device of the present invention enables rapid sample preparation at constant temperature and under sterilization, and a biological tissue in which cells are kept morphologically sufficiently stable for relatively long-term culture. Thin sliced samples can be easily and reliably produced. This technology will be very useful for drug research and development, medical diagnosis and treatment.

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Abstract

L'invention concerne une méthode de préparation d'un échantillon de tissu biologique en tranches dans lequel les cellules sont conservées morphologiquement stables, un instrument et un dispositif afférent. Une combinaison d'au moins deux couteaux lamellaires, en céramique, ayant les mêmes bords linéaires ou courbés, et maintenus parallèles l'un à l'autre à des intervalles spécifiés avec leurs bords également maintenus parallèles mutuellement, est utilisée pour découper en tranches un bloc de tissu biologique en une épaisseur prédéfinie afin d'obtenir des tranches de tissu biologique, et des éléments de pression, présentant chacun une épaisseur spécifiée, pouvant être insérée entre les couteaux sont insérés respectivement dans les espaces entre les couteaux afin d'expulser les tranches de tissu biologique d'entre les couteaux.
PCT/JP2000/003107 1999-10-07 2000-05-15 Methode de preparation d'un echantillon de tissu biologique et instrument et dispositif afferent WO2001027586A1 (fr)

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Cited By (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2002088668A1 (fr) * 2001-04-16 2002-11-07 Institute Of Whole Body Metabolism Dispositif de fabrication d'une tranche fine de tissu biologique
EP1331479A1 (fr) * 2000-10-06 2003-07-30 Institute of Whole Body Metabolism Procede d'imagerie quantifie a deux dimensions pour distinguer et quantifier une excroissance tissulaire ou similaire
JP2004121064A (ja) * 2002-10-01 2004-04-22 Jfe Steel Kk 遺伝子制御用のセラミック被覆針
JP2005533102A (ja) * 2002-07-12 2005-11-04 イッサム リサーチ ディベロップメント カンパニー オブ ザ ヘブリュー ユニバーシティー オブ エルサレム 微小器官によって生物学的プロセスを誘発する方法と装置
JP2006145539A (ja) * 2004-11-20 2006-06-08 Leica Microsystems Nussloch Gmbh 滅菌装置を有するクライオスタット
WO2008053916A1 (fr) * 2006-11-01 2008-05-08 Kurume University Dispositif de guidage de coupe pour préparer une section de texture, dispositif de préparation de section de texture, et procédé pour préparer une section de texture
JP2008222640A (ja) * 2007-03-13 2008-09-25 Kitazato Supply:Co Ltd 採取組織片ガラス化凍結保存用具、組織片凍結保存用プレートおよび採取組織片ガラス化凍結保存方法
US7651507B2 (en) 2003-03-03 2010-01-26 Kci Licensing, Inc. Tissue processing system
CN104792571A (zh) * 2014-01-17 2015-07-22 杨涛 乳腺癌保乳标本取量台
WO2015115439A1 (fr) * 2014-01-29 2015-08-06 株式会社ジェイ・エム・エス Dispositif de découpe de tissu biologique et son utilisation
US9127084B2 (en) 2006-09-14 2015-09-08 Medgenics Medical Israel Ltd. Long lasting drug formulations
US9155749B2 (en) 2006-09-14 2015-10-13 Medgenics Medical Israel Ltd. Long lasting drug formulations
CN106289856A (zh) * 2016-08-22 2017-01-04 山东大学齐鲁医院 一种大截面及大厚度脑组织切片的制作装置及方法
WO2017009800A1 (fr) 2015-07-16 2017-01-19 Ecole Polytechnique Federale De Lausanne (Epfl) Dispositif de découpe de tranches et de caractérisation d'échantillon à haut débit
WO2019208403A1 (fr) * 2018-04-25 2019-10-31 シャープ株式会社 Gabarit
JP2020063950A (ja) * 2018-10-16 2020-04-23 株式会社ディスコ 肉片切断方法

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* Cited by examiner, † Cited by third party
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JP4510908B2 (ja) * 2008-05-30 2010-07-28 周 市原 摘出物スライス補助装置及び摘出物スライス補助セット
JP6238397B2 (ja) * 2013-08-01 2017-11-29 国立大学法人 香川大学 細胞空間分画装置および微細構造刃
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Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS57166154U (fr) * 1981-04-15 1982-10-20
JPS6066494U (ja) * 1983-10-07 1985-05-11 有限会社太田精機製作所 封書の開封機構を備えたカツタ−
EP0194652A1 (fr) * 1985-03-13 1986-09-17 Shin-Etsu Chemical Co., Ltd. Lame revêtue pour un microtome et procédé pour sa fabrication
US4723420A (en) * 1984-07-12 1988-02-09 C. Reichert Optische Werke A.G. Apparatus for treating specimens at low temperature
JPH01104157A (ja) * 1987-07-07 1989-04-21 Mitsui Petrochem Ind Ltd 培養装置
JPH01285185A (ja) * 1988-05-10 1989-11-16 Taisei Corp 環境調節施設
US5070935A (en) * 1986-10-14 1991-12-10 Hellmuth Sitte Refrigerated chamber for obtaining thin slices at low temperature
JPH055682A (ja) * 1991-06-28 1993-01-14 Jeol Ltd ミクロトーム用ナイフ親水化処理方法及び装置
JPH07218398A (ja) * 1994-01-31 1995-08-18 Seitai Kagaku Kenkyusho:Kk 生物組織試料作成方法および器具

Patent Citations (9)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JPS57166154U (fr) * 1981-04-15 1982-10-20
JPS6066494U (ja) * 1983-10-07 1985-05-11 有限会社太田精機製作所 封書の開封機構を備えたカツタ−
US4723420A (en) * 1984-07-12 1988-02-09 C. Reichert Optische Werke A.G. Apparatus for treating specimens at low temperature
EP0194652A1 (fr) * 1985-03-13 1986-09-17 Shin-Etsu Chemical Co., Ltd. Lame revêtue pour un microtome et procédé pour sa fabrication
US5070935A (en) * 1986-10-14 1991-12-10 Hellmuth Sitte Refrigerated chamber for obtaining thin slices at low temperature
JPH01104157A (ja) * 1987-07-07 1989-04-21 Mitsui Petrochem Ind Ltd 培養装置
JPH01285185A (ja) * 1988-05-10 1989-11-16 Taisei Corp 環境調節施設
JPH055682A (ja) * 1991-06-28 1993-01-14 Jeol Ltd ミクロトーム用ナイフ親水化処理方法及び装置
JPH07218398A (ja) * 1994-01-31 1995-08-18 Seitai Kagaku Kenkyusho:Kk 生物組織試料作成方法および器具

Cited By (23)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP1331479A1 (fr) * 2000-10-06 2003-07-30 Institute of Whole Body Metabolism Procede d'imagerie quantifie a deux dimensions pour distinguer et quantifier une excroissance tissulaire ou similaire
EP1331479A4 (fr) * 2000-10-06 2005-01-12 Inst Of Whole Body Metabolism Procede d'imagerie quantifie a deux dimensions pour distinguer et quantifier une excroissance tissulaire ou similaire
WO2002088668A1 (fr) * 2001-04-16 2002-11-07 Institute Of Whole Body Metabolism Dispositif de fabrication d'une tranche fine de tissu biologique
JP2005533102A (ja) * 2002-07-12 2005-11-04 イッサム リサーチ ディベロップメント カンパニー オブ ザ ヘブリュー ユニバーシティー オブ エルサレム 微小器官によって生物学的プロセスを誘発する方法と装置
JP2004121064A (ja) * 2002-10-01 2004-04-22 Jfe Steel Kk 遺伝子制御用のセラミック被覆針
US7651507B2 (en) 2003-03-03 2010-01-26 Kci Licensing, Inc. Tissue processing system
US8162957B2 (en) 2003-03-03 2012-04-24 Kci Licensing, Inc. Tissue processing system
JP2006145539A (ja) * 2004-11-20 2006-06-08 Leica Microsystems Nussloch Gmbh 滅菌装置を有するクライオスタット
US9127084B2 (en) 2006-09-14 2015-09-08 Medgenics Medical Israel Ltd. Long lasting drug formulations
US9155749B2 (en) 2006-09-14 2015-10-13 Medgenics Medical Israel Ltd. Long lasting drug formulations
US8122804B2 (en) 2006-11-01 2012-02-28 Kurume University Slicing guide device for preparing texture slices, texture slices preparing device, and method for preparing texture slices
WO2008053916A1 (fr) * 2006-11-01 2008-05-08 Kurume University Dispositif de guidage de coupe pour préparer une section de texture, dispositif de préparation de section de texture, et procédé pour préparer une section de texture
JP2008222640A (ja) * 2007-03-13 2008-09-25 Kitazato Supply:Co Ltd 採取組織片ガラス化凍結保存用具、組織片凍結保存用プレートおよび採取組織片ガラス化凍結保存方法
CN104792571A (zh) * 2014-01-17 2015-07-22 杨涛 乳腺癌保乳标本取量台
CN104792571B (zh) * 2014-01-17 2017-07-11 杨涛 乳腺癌保乳标本取量台
WO2015115439A1 (fr) * 2014-01-29 2015-08-06 株式会社ジェイ・エム・エス Dispositif de découpe de tissu biologique et son utilisation
JPWO2015115439A1 (ja) * 2014-01-29 2017-03-23 株式会社ジェイ・エム・エス 生体組織切断装置およびその用途
US10036689B2 (en) 2014-01-29 2018-07-31 Jms Co., Ltd. Biological tissue cutting device and use thereof
WO2017009800A1 (fr) 2015-07-16 2017-01-19 Ecole Polytechnique Federale De Lausanne (Epfl) Dispositif de découpe de tranches et de caractérisation d'échantillon à haut débit
CN106289856A (zh) * 2016-08-22 2017-01-04 山东大学齐鲁医院 一种大截面及大厚度脑组织切片的制作装置及方法
CN106289856B (zh) * 2016-08-22 2019-04-30 山东大学齐鲁医院 一种大截面及大厚度脑组织切片的制作装置及方法
WO2019208403A1 (fr) * 2018-04-25 2019-10-31 シャープ株式会社 Gabarit
JP2020063950A (ja) * 2018-10-16 2020-04-23 株式会社ディスコ 肉片切断方法

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